Toxins produced by Clostridium botulinum are classified into types A to G depending on the difference in serotypes. The toxin types A, B, E, and F have a serious influence on the human health. There are four kinds of botulinum diseases in humans, such as “foodborne botulism (botulinum food poisoning)”, “infant botulism”, “wound botulism” and “infectious botulism”. The toxin type-A is involved in 26% of botulinum food poisoning. The toxin type-A is involved in 26% of botulinum food poisoning. The toxin types B and E are followed by this, and toxin type-F is relatively low in the ratio involved in botulinum food poisoning. In addition, it is reported that wound botulism is caused only by toxin type-A or type-B (Non-Patent Document 1).
Food poisoning botulism in adults is usually developed by oral intake of toxins produced as a result of the growth of bacteria and spores in food, while in the case of infant botulism, symptoms of poisoning are caused by the bacterial growth and proliferation leading to toxin production in the intestinal tract as a result of oral intake of bacteria and spores in food such as honey and the like. Although toxins that cause food poisoning differ somewhat depending on countries, cases caused by toxin types A, B, and E have been reported. Moreover, the majority of the infant botulisms are type-A and type B, but toxin types E and F by C. butiricum also have been reported.
Botulinum toxin is a molecule having a structure in which an S—S bond (disulfide bond) is formed between the heavy chain and the light chain. The botulinum toxin blocks the neurotransmission to muscles, causing hereby flaccid paralysis. When botulinum toxin comes into the respiratory tract or respiratory muscle, fatal breathing disorder will be caused. The mortality rate by the botulinum toxin is 12%, and it becomes higher in a specific risk group (Non-Patent Document 3).
On the other hand, since the botulinum toxin is the most fatal toxin as a biological toxin (the fatal dose of the purified toxin type-A in a human of 70 kg is 0.09-0.15 μg (i.v. or i.m.), 0.70-0.90 μg (inhalation), and 70 μg (even oral) (JAMA. 2001 Feb. 28; 285(8):1059-70)), such a botulinum toxin possesses a particularity such as an applicability to the bioterrorism as an another aspect other than usual poisoning. Therefore, a social attention has been paid to it.
The neutralizing agent for botulinum toxins, approved in Japan, is a preparation using an equine serum as a raw material. This preparation may have a danger to cause an immunoreaction such as anaphylactic reaction, etc. because it contains, as a main component, globulin derived from equine serum, i.e. a heterologous protein to humans. In addition, there are problems of unknown viral infections and serum diseases in the preparation. Furthermore, stable supply of this preparation is also difficult. Besides, since one year or more are needed for the production of the preparations, emergency response to bioterrorism and the like is in a difficult situation. In addition, this preparation is not suitable for a storage purpose because there is a problem in the management of horse rearing.
As for infant botulism, it is a current state that treatment with this equine globulin preparation has not been employed because of avoidance of serious side effects such as anaphylactic reactions and the like. In the United States of America, a specific treatment for the infant botulism is performed with use of a human globulin preparation prepared from the plasma having a high neutralizing antibody titer obtained by immunizing a healthy person with a botulinum toxin. Actually, in the United States of America, the drug under the brand name called BabyBIG® for the infant botulism has been granted orphan drug designation for its marketing by FDA. However, this method holds problems of raw material availability and biohazard issues in addition to ethical issues, as well as problems of various virus inspections to be needed from the viewpoint of safe security necessary in the production. In addition, the preparation has not yet been approved in Japan and cannot be used.
It was reported to have succeeded in producing a recombinant neutralizing chimeric antibody against toxin type-A (Patent Document 1). However, it is necessary to solve the problems such as appearance of HACA (human anti-chimeric antibody) or anaphylactic reactions when the antibody is used.
Meanwhile, a research and development for toxin neutralization have been performed from the viewpoint of preparing for particularly bioterrorism threats in the United States of America, and a method of producing a phage antibody library from human B lymphocytes obtained by immunization with a botulinum toxin pentamer and selecting a desired full length human neutralizing antibody from the library was undertaken. However, such a method has not succeeded in obtaining an antibody clone that exerts a sufficient neutralizing activity against the toxin type-A alone. Therefore, it could not but enhance the neutralizing activity by mixing (oligo cloning) one kind of human antibody obtained as a result of phage display screening, with one kind of clone derived from XENO mice and further a humanized mouse antibody obtained by immunization in mice. Accordingly, intended purpose, that is, neutralization of the toxin with the full length human neutralizing antibody has not been achieved (Patent Documents 2 and 3, and Non-Patent Documents 4, 5, and 6).    Patent Document 1: Japanese Patent Laid-Open Publication No. 2006-311857    Patent Document 2: PCT WO 2005/016232 Pamphlet    Patent Document 3: PCT WO 2007/094754 Pamphlet    Non-Patent Document 1: H. Sugiyama, Microbiol. Rev. 44:419, 1980    Non-Patent Document 2: S. Arnon, Epidemiol. Rev. 3:45, 1981    Non-Patent Document 3: C. O. Tacket et al., Am. J. Med. 76:794, 1984    Non-Patent Document 4: P. Amersdorfer et al., Infect. Immun. 65(9):3743, 1997    Non-Patent Document 5: P. Amersdorfer et al., Vaccine 20: 1640, 2002    Non-Patent Document 6: Nowakowski et al., Proc. Natl. Acad. Sci. USA 99(17):11346, 2002